Extensive preclinical studies, as well as results obtained from clinical trials, suggest that vaccination with an anti-idiotype (Id) antibody (3H1) that mimics an epitope of human carcinoembryonic antigen (CEA) has the potential to augment survival benefits. Anti-ld 3H1 breaks immune tolerance to CEA and induces anti-CEA antibody as well as CD4+T helper (Th1) responses in colorectal cancer patients and also in mice transgenic for CEA. This anti-ld approach in its current form, although promising, will need improvements to realize its full potential. Suitable murine tumor models will be used to explore strategies that will significantly improve the therapeutic impact of this vaccine. The proposal is based on the hypothesis that stimulating a CEA-specific CD4+ T cell response, ie T-help, in the host, will provide critical help for priming and activation of CEA-specific CTL, the major effector cells (CD8+ T cells) for tumor destruction. We hypothesize that the combination of CD4+ with CD8+ T cell epitopes will further augment the anti-tumor immune responses. The specific aims of this proposal are: 1) to determine whether vaccination with 3H1, which will generate anti-CEA Ab and T-help, in combination with mRNA derived from CEA, using dendritic cells (DC) as APC, will induce CTL and engender therapeutic immunity in an established tumor model in C57BL/6 mice (H2kb), double transgenic for human CEA and HLA-A2; 2) to explore whether a combination of 3H1 with HLA-A2 restricted known agonist CTL epitopes of CEA, using DC as APC, will work better in the above tumor model; 3) to test whether the idio-peptides, (LCD-2 and CEA-B) derived from the structure of 3H1 and CEA based on the amino acid sequence homology, which also induce CD4+Th1 help will be more immunogenic in combination with the agonist CTL peptides of CEA. The criteria for selection of the optimal regimen for vaccination will be based on the ability to invoke anti-tumor activities in vitro and in vivo. We will measure the antibody titer, in vitro CTL activity, intra-cellular cytokine levels and in vivo tumor regression. In Aim 4, we will further explore methods to boost tumor-specific CD4+ as well as CD8+ T cell responses in vivo by coadministration of agents such as IL-2, IL-12, CpG ODN or anti-CTLA4 antibody. We will test for any possible autoimmune responses in CEA-expressing normal organs of mice by histopathological analysis (Aim 5). Promising strategies indicated by these studies will be finally evaluated in Aim 6 in the murine Apc knock-out transgenic mice expressing CEA and HLA-A2, which arguably, are the best murine model for colon cancer, as it closely resembles the human disease. The results obtained from these studies will help design improved therapeutic vaccines for the treatment of CEA+ tumors and can be incorporated readily into our ongoing clinical programs.